Lysosomal membrane protein Sidt2 knockout induces apoptosis of human hepatocytes in vitro independent of the autophagy-lysosomal pathway.
10.12122/j.issn.1673-4254.2023.04.18
- Author:
Jiating XU
1
;
Mengya GENG
1
;
Haijun LIU
2
;
Wenjun PEI
2
;
Jing GU
1
;
Mengxiang QI
3
;
Yao ZHANG
2
;
Kun LÜ
4
;
Yingying SONG
1
;
Miaomiao LIU
2
;
Xin HU
1
;
Cui YU
5
;
Chunling HE
1
;
Lizhuo WANG
2
;
Jialin GAO
1
Author Information
1. Department of Endocrinology and Genetic Metabolism, Yijishan Hospital of Wannan Medical College, Wuhu 241002, China.
2. Anhui Provincial Key Laboratory of Biological Macro-molecules Research, Yijishan Hospital of Wannan Medical College, Wuhu 241002, China.
3. School of Clinical Medicine, Yijishan Hospital of Wannan Medical College, Wuhu 241002, China.
4. Central Laboratory, Yijishan Hospital of Wannan Medical College, Wuhu 241002, China.
5. Department of Endocrinology, Second Affiliated Hospital of Wannan Medical College, Wuhu 241002, China.
- Publication Type:Journal Article
- Keywords:
Sidt2;
apoptosis;
autophagy;
human liver cells;
proliferation
- MeSH:
Humans;
Lysosome-Associated Membrane Glycoproteins/metabolism*;
Autophagy;
Apoptosis;
Hepatocytes;
Lysosomes/metabolism*;
Chloroquine/pharmacology*;
Nucleotide Transport Proteins/metabolism*
- From:
Journal of Southern Medical University
2023;43(4):637-643
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVE:To explore the regulatory mechanism of human hepatocyte apoptosis induced by lysosomal membrane protein Sidt2 knockout.
METHODS:The Sidt2 knockout (Sidt2-/-) cell model was constructed in human hepatocyte HL7702 cells using Crispr-Cas9 technology.The protein levels of Sidt2 and key autophagy proteins LC3-II/I and P62 in the cell model were detected using Western blotting, and the formation of autophagosomes was observed with MDC staining.EdU incorporation assay and flow cytometry were performed to observe the effect of Sidt2 knockout on cell proliferation and apoptosis.The effect of chloroquine at the saturating concentration on autophagic flux, proliferation and apoptosis of Sidt2 knockout cells were observed.
RESULTS:Sidt2-/- HL7702 cells were successfully constructed.Sidt2 knockout significantly inhibited the proliferation and increased apoptosis of the cells, causing also increased protein expressions of LC3-II/I and P62(P < 0.05) and increased number of autophagosomes.Autophagy of the cells reached a saturated state following treatment with 50 μmol/L chloroquine, and at this concentration, chloroquine significantly increased the expressions of LC3B and P62 in Sidt2-/- HL7702 cells.
CONCLUSION:Sidt2 gene knockout causes dysregulation of the autophagy pathway and induces apoptosis of HL7702 cells, and the latter effect is not mediated by inhibiting the autophagy-lysosomal pathway.
